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As clinically demonstrated by the success of immunotherapies to improve survival outcomes, tumors are known to gain a survival advantage by circumventing immune surveillance. A defining feature of this is the creation and maintenance of a tumor immune microenvironment (TIME) that directly and indirectly alters the host’s immunologic signaling pathways through a variety of mechanisms. Tumor-intrinsic mechanisms that instruct the formation and maintenance of the TIME have been an area of intensive study, such as the identification and characterization of soluble factors actively and passively released by tumor cells that modulate immune cell function. In particular, damage-associated molecular pattern (DAMP) molecules typically released by necrotic tumor cells are recognized by innate immune receptors such as Toll-like receptors (TLRs) and stimulate immune cells within TIME. Given their broad and potent effects on the immune system, a better understanding for how DAMP and TLR interactions sculpt the TIME to favor tumor growth would identify new strategies and approaches for cancer immunotherapy.

One of most challenging issues in tumor immunology is a better understanding of the dynamics in the accumulation of myeloid-derived suppressor cells (MDSCs) in the tumor microenvironment (TIME), as this would lead to the development of new cancer therapeutics. Here, we show that translationally controlled tumor protein (TCTP) released by dying tumor cells is an immunomodulator crucial to full-blown MDSC accumulation in the TIME. We provide evidence that extracellular TCTP mediates recruitment of the polymorphonuclear MDSC (PMN-MDSC) population in the TIME via activation of Toll-like receptor-2. As further proof of principle, we show that inhibition of TCTP suppresses PMN-MDSC accumulation and tumor growth. In human cancers, we find an elevation of TCTP and an inverse correlation of TCTP gene dosage with antitumor immune signatures and clinical prognosis. This study reveals the hitherto poorly understood mechanism of the MDSC dynamics in the TIME, offering a new rationale for cancer immunotherapy. Cell death in the context of cancer therapies is often associated with immunogenicity. Taniguchi and colleagues instead find that release of the cellular protein TCTP following cell death triggers an immunosuppressive pathway in the tumor microenvironment.

The transcription factor IRF5 has been implicated as a therapeutic target for the autoimmune disease systemic lupus erythematosus (SLE). However, IRF5 activation status during the disease course and the effects of IRF5 inhibition after disease onset are unclear. Here, we show that SLE patients in both the active and remission phase have aberrant activation of IRF5 and interferon-stimulated genes. Partial inhibition of IRF5 is superior to full inhibition of type I interferon signaling in suppressing disease in a mouse model of SLE, possibly due to the function of IRF5 in oxidative phosphorylation. We further demonstrate that inhibition of IRF5 via conditional Irf5 deletion and a newly developed small-molecule inhibitor of IRF5 after disease onset suppresses disease progression and is effective for maintenance of remission in mice. These results suggest that IRF5 inhibition might overcome the limitations of current SLE therapies, thus promoting drug discovery research on IRF5 inhibitors., IRF5 is a potential target for therapy in systemic lupus erythematosus (SLE). Here the authors show using mouse SLE-like models that genetic or chemical inhibition of IRF5 after SLE onset could be more effective than, or an add on for, currently utilised type I interferon inhibition.

The signal‐transducing innate receptors represent classes of pattern recognition receptors (PRRs) that play crucial roles in the first line of the host defense against infections by the recognition of pathogen‐derived molecules. Because of their poorly discriminative nature compared with antigen receptors of the adaptive immune system, they also recognize endogenous molecules and evoke immune responses without infection, resulting in the regulation of tumor immunity. Therefore, PRRs may be promising targets for effective cancer immunotherapy, either by activating or inhibiting them. Here, we summarize our current knowledge of signal‐transducing PRRs in the regulation of tumor immunity., The signal transducing innate receptors represent classes of pattern recognition receptors (PRRs) that play crucial roles in the first line of host defense against infections by the recognition of pathogen‐derived molecules. They also recognize endogenous molecules and evoke immune responses without infection, resulting in the regulation of tumor immunity. Hence, PRRs may be the promising targets for effective cancer immunotherapy. Here, we summarize our current knowledge of signal‐transducing PRRs in the regulation of tumor immunity.

Dying or damaged cells that are not completely eradicated by the immune system release their intracellular components in the extracellular space. Aberrant exposure of the damage-associated molecules to the immune system is often associated with inflammation and cancer pathogenesis. Thus, elucidating the role of damage-associated molecules in inducing sterile immune responses is crucial. In this study, we show that prostaglandin E2 (PGE2) is produced in the supernatants from several types of canine necrotic tumor cell lines. Inhibition of PGE2 production by indomethacin, a potent inhibitor of cyclooxygenase (COX) enzymes, induces the expression of tumor necrosis factor (Tnf) mRNA in the necrotic tumor cell supernatants. These results comply with the previous observations reported in mouse cell lines. Furthermore, comprehensive ribonucleic acid-sequencing (RNA-seq) analysis revealed that three categories of genes were induced by the damage-associated molecules: (i) a group of PGE2-inducible genes, (ii) genes that promote inflammation and are suppressed by PGE2, and (iii) a group of genes not suppressed by PGE2. Collectively, our findings reveal the hitherto unknown immune regulatory system by PGE2 and damage-associated molecules, which may have clinical implications in inflammation and cancer.

Significance Immunogenic damage-associated molecular patterns (DAMPs), typically released from dying cells, can evoke chronic inflammatory or autoimmune disorders via their activation of innate immune receptors. Since an association of RNA-sensing Toll-like receptor 7 (TLR7) signaling with autoimmunity is well-documented, identification of a DAMP(s) that triggers TLR7 is critical to understanding the disease pathogenesis. By generating the synthetic compound KN69 that inhibits autoimmunity in mouse models, we identified U11 small nuclear RNA (U11snRNA) as a target of KN69 and strong activator of TLR7. We found a correlation between high serum level of U11snRNA and autoimmune diseases in human subjects and mouse models. Finally, we generated TLR7 agonists and TLR7 antagonists. Our study provides therapeutic insight into autoimmunity and other diseases., The activation of innate immune receptors by pathogen-associated molecular patterns (PAMPs) is central to host defense against infections. On the other hand, these receptors are also activated by immunogenic damage-associated molecular patterns (DAMPs), typically released from dying cells, and the activation can evoke chronic inflammatory or autoimmune disorders. One of the best known receptors involved in the immune pathogenesis is Toll-like receptor 7 (TLR7), which recognizes RNA with single-stranded structure. However, the causative DAMP RNA(s) in the pathogenesis has yet to be identified. Here, we first developed a chemical compound, termed KN69, that suppresses autoimmunity in several established mouse models. A subsequent search for KN69-binding partners led to the identification of U11 small nuclear RNA (U11snRNA) as a candidate DAMP RNA involved in TLR7-induced autoimmunity. We then showed that U11snRNA robustly activated the TLR7 pathway in vitro and induced arthritis disease in vivo. We also found a correlation between high serum level of U11snRNA and autoimmune diseases in human subjects and established mouse models. Finally, by revealing the structural basis for U11snRNA’s ability to activate TLR7, we developed more potent TLR7 agonists and TLR7 antagonists, which may offer new therapeutic approaches for autoimmunity or other immune-driven diseases. Thus, our study has revealed a hitherto unknown immune function of U11snRNA, providing insight into TLR7-mediated autoimmunity and its potential for further therapeutic applications.

Increase of the enteric bacteriophages (phage), components of the enteric virome, has been associated with the development of inflammatory bowel diseases. However, little is known about how a given phage contributes to the regulation of intestinal inflammation. In this study, we isolated a new phage associated with Enterococcus gallinarum, named phiEG37k, the level of which was increased in C57BL/6 mice with colitis development. We found that, irrespective of the state of inflammation, over 95% of the E. gallinarum population in the mice contained phiEG37k prophage within their genome and the phiEG37k titers were proportional to that of E. gallinarum in the gut. To explore whether phiEG37k impacts intestinal homeostasis and/or inflammation, we generated mice colonized either with E. gallinarum with or without the prophage phiEG37k. We found that the mice colonized with the bacteria with phiEG37k produced more Mucin 2 (MUC2) that serves to protect the intestinal epithelium, as compared to those colonized with the phage-free bacteria. Consistently, the former mice were less sensitive to experimental colitis than the latter mice. These results suggest that the newly isolated phage has the potential to protect the host by strengthening mucosal integrity. Our study may have clinical implication in further understanding of how bacteriophages contribute to the gut homeostasis and pathogenesis.

Dysregulation of inflammatory cytokines in keratinocytes promote the pathogenesis of the skin inflammation, such as allergic contact dermatitis (ACD). High-mobility group box 1 protein (HMGB1) has been implicated in the promotion of skin inflammation upon its extracellular release as a damage-associated molecular pattern molecule. However, whether and how HMGB1 in keratinocytes contributes to ACD and other skin disorders remain elusive. In this study, we generated conditional knockout mice in which the Hmgb1 gene is specifically deleted in keratinocytes, and examined its role in ACD models. Interestingly, the mutant mice showed exacerbated skin inflammation, accompanied by increased ear thickening in 2,4-dinitrofluorobenezene-induced ACDs. The mRNA expression of interleukin-24 (IL-24), a cytokine known to critically contribute to ACD pathogenesis, was elevated in skin lesions of the mutant mice. As with constitutively expressed, IL-4–induced Il24 mRNA, expression was also augmented in the Hmgb1-deficient keratinocytes, which would account for the exacerbation of ACD in the mutant mice. Mechanistically, we observed an increased binding of trimethyl histone H3 (lys4) (H3K4me3), a hallmark of transcriptionally active genes, to the promoter region of the Il24 gene in the hmgb1-deficient cells. Thus, the nuclear HMGB1 is a critical “gate keeper” in that the dermal homeostasis is contingent to its function in chromatin remodeling. Our study revealed a facet of nuclear HMGB1, namely its antiinflammatory function in keratinocytes for the skin homeostasis.

The commensal microbiota within the gastrointestinal tract is essential in maintaining homeostasis. Indeed, dysregulation in the repertoire of microbiota can result in the development of intestinal immune-inflammatory diseases. Further, this immune regulation by gut microbiota is important systemically, impacting health and disease of organ systems beyond the local environment of the gut. What has not been explored is how distant organs might in turn shape the microbiota via microbe-targeted molecules. Here, we provide evidence that surfactant protein D (SP-D) synthesized in the gallbladder and delivered into intestinal lumen binds selectively to species of gut commensal bacteria. SP-D-deficient mice manifest intestinal dysbiosis and show a susceptibility to dextran sulfate sodium-induced colitis. Further, fecal transfer from SP-D-deficient mice to wild-type, germ-free mice conveyed colitis susceptibility. Interestingly, colitis caused a notable increase in Sftpd gene expression in the gallbladder, but not in the lung, via the activity of glucocorticoids produced in the liver. These findings describe a unique mechanism of interorgan regulation of intestinal immune homeostasis by SP-D with potential clinical implications such as cholecystectomy.

Malignant ascites manifests as an end-stage event during the progression of a number of cancers and lacks a generally accepted standard therapy. Interferon-β (IFN-β) has been used to treat several cancer indications; however, little is known about the efficacy of IFN-β on malignant ascites. In the present study, we report on the development of a novel, engineered form of human and murine IFN-β, each conjugated with a polyethylene glycol molecule (PEG-hIFN-β and PEG-mIFN-β, respectively). We provide evidence that these IFN-β molecules retain anti-viral potency comparable to unmodified IFN-β in vitro and manifested improved pharmacokinetics in vivo. Interestingly, PEG-mIFN-β significantly inhibited the accumulation of ascites fluid and vascular permeability of the peritoneal membrane in models of ovarian cancer and gastric cancer cell xenograft mice. We further show that PEG-hIFN-β directly suppresses VEGF165 -induced hyperpermeability in a monolayer of human vascular endothelial cells and that PEG-mIFN-β enhanced gene expression for a number of cell adhesion related molecules in mouse vascular endothelial cells. Taken together, these findings unveil a hitherto unrecognized potential of IFN-β in maintaining vascular integrity, and provide proof-of-mechanism for a novel and long-acting pegylated hIFN-β for the therapeutic treatment of malignant ascites.

Cellular components released into the external milieu as a result of cell death and sensed by the body are generally termed damage-associated molecular patterns (DAMPs). Although DAMPs are conventionally thought to be protective to the host by evoking inflammatory responses important for immunity and wound repair, there is the prevailing notion that dysregulated release of DAMPs can also underlie or exacerbate disease development. However, the critical issue for how resultant DAMP-mediated responses are regulated has heretofore not been fully addressed. In the present study, we identify prostaglandin E2 (PGE2) as a DAMP that negatively regulates immune responses. We show that the production of PGE2 is augmented under cell death-inducing conditions via the transcriptional induction of the cyclooxygenase 2 (COX2) gene and that cell-released PGE2 suppresses the expression of genes associated with inflammation, thereby limiting the cell's immunostimulatory activities. Consistent with this, inhibition of the PGE2 synthesis pathway potentiates the inflammation induced by dying cells. We also provide in vivo evidence for a protective role of PGE2 released upon acetaminophen-induced liver injury as well as a pathogenic role for PGE2 during tumor cell growth. Our study places this classically known lipid mediator in an unprecedented context-that is, an inhibitory DAMP vis-à-vis activating DAMPs, which may have translational implications for designing more effective therapeutic regimens for inflammation-associated diseases.

Significance IFN regulatory factor 3 (IRF3) is one of the most well-characterized transcription factors involved in the regulation of innate immune responses. Many studies have used conventional Irf3 -deficient mice to analyze the functions of IRF3 in immunity and other biological systems; however, these mice were found to carry another null mutation in the Bcl2l12 gene, which has confounded the analyses of these mice. For this study, we generated conditional Irf3 -deficient mice while leaving the Bcl2l12 gene intact. Conditional ablation of IRF3 in myeloid cells showed increased survival when challenged with endotoxin-induced shock, indicating a myeloid cell-specific role of IRF3. The IRF3 conditional knockout mouse may be a useful tool to identify cell type-specific functions of this transcription factor.

Recent years have seen a number of regulatory approvals for immune oncology or immunotherapies based on their ability to enhance antitumor immune responses. Nevertheless, the majority of patients remain refractory to these treatments; hence, new therapies that augment current immunotherapies are required. Innate immune receptors that recognize nucleic acids are potent activators of subsequent T‐cell responses and, as a result, can evoke potent antitumor immune responses. Herein, we present a novel compound N‐{3‐[(1,4′‐bipiperidin)‐1′‐yl]propyl}‐6‐[4‐(4‐methylpiperazin‐1‐yl)phenyl]picolinamide (SINCRO; STING‐mediated interferon‐inducing and cytotoxic reagent, original) as an anticancer drug that activates the cytosolic DNA‐sensing STING (stimulator of interferon genes) signaling pathway leading to the induction of type I interferon (IFN) genes. Indeed, IFN‐β gene induction by SINCRO is abolished in STING‐deficient cells. In addition to its IFN‐inducing activity, SINCRO shows STING‐independent cytotoxic activity against cancer cells. SINCRO does not evoke DNA double‐strand break or caspase‐3 cleavage. Thus, SINCRO induces cell death in a method different from conventional apoptosis‐inducing pathways. Finally, we provide evidence that giving SINCRO significantly attenuates in vivo tumor growth by both type I IFN‐dependent and independent mechanisms. Thus, SINCRO is an attractive anticancer compound with dual function in that it evokes type I IFN response to promote antitumor immunity as well as inducing tumor cell death. SINCRO may provide a new platform for the development of drugs for effective cancer therapy.

The regulation of intestinal homeostasis by the immune system involves the dynamic interplay between gut commensal microbiota and resident immune cells. It is well known that a large and diverse lymphocyte antigen receptor repertoire enables the immune system to recognize and respond to a wide range of invading pathogens. There is also an emerging appreciation for a critical role the T-cell receptor (TCR) repertoire serves in the maintenance of peripheral tolerance by regulatory T cells (Tregs). Nevertheless, how the diversity of the TCR repertoire in Tregs affects intestinal homeostasis remains unknown. To address this question, we studied mice whose T cells express a restricted TCR repertoire. We observed the development of spontaneous colitis, accompanied by the induction of T-helper type 17 cells in the colon that is driven by gut commensal microbiota. We provide further evidence that a restricted TCR repertoire causes a loss of tolerogenicity to microbiota, accompanied by a paucity of peripherally derived, Helios(-) Tregs and hyperactivation of migratory dendritic cells. These results thus reveal a new facet of the TCR repertoire in which Tregs require a diverse TCR repitoire for intestinal homeostasis, suggesting an additional driving force in the evolutional significance of the TCR repertoire.

Interferons (IFNs) are a broad class of cytokines elicited on challenge to the host defense and are essential for mobilizing immune responses to pathogens. Divided into three classes, type I, type II, and type III, all IFNs share in common the ability to evoke antiviral activities initiated by the interaction with their cognate receptors. The nine-member IFN regulatory factor (IRF) family, first discovered in the context of transcriptional regulation of type I IFN genes following viral infection, are pivotal for the regulation of the IFN responses. In this review, we briefly describe cardinal features of the three types of IFNs and then focus on the role of the IRF family members in the regulation of each IFN system.

High-mobility group box 1 (HMGB1) was first described as an architectural chromatin-binding protein. Today, a wealth of evidence indicates that this protein is very versatile and serves an amazing assortment of roles in the nucleus, cytoplasm and extracellular milieu. As a result, HMGB1 is fast becoming recognized as a key regulator of protective and pathological immune responses. Whilst acknowledging the many functions of HMGB1 and its family members, we focus this review on their role as broad effectors of immune responses mediated by nucleic acids. In addition, we touch upon the recent progress in determining the in vivo role of HMGB1 as revealed by the study of mice conditionally null for the Hmgb1 gene.

High-mobility group box 1 (HMGB1) is a DNA-binding protein abundantly expressed in the nucleus that has gained much attention for its regulation of immunity and inflammation. Despite this, whether and how HMGB1 contributes to protective and/or pathological responses in vivo is unclear. In this study, we constructed Hmgb1-floxed (Hmgb1(f)(/f)) mice to achieve the conditional inactivation of the gene in a cell- and tissue-specific manner by crossing these mice with an appropriate Cre recombinase transgenic strain. Interestingly, although mice with HMGB1 ablation in myeloid cells apparently develop normally, they are more sensitive to endotoxin shock compared with control mice, which is accompanied by massive macrophage cell death. Furthermore, these mice also show an increased sensitivity to Listeria monocytogenes infection. We also provide evidence that the loss of HMGB1 in macrophages results in the suppression of autophagy, which is commonly induced by lipopolysaccharide stimulation or L. monocytogenes infection. Thus, intracellular HMGB1 contributes to the protection of mice from endotoxemia and bacterial infection by mediating autophagy in macrophages. These newly generated HMGB1 conditional knockout mice will serve a useful tool with which to study further the in vivo role of this protein in various pathological conditions.

A major function of innate immune receptors is to recognize pathogen-associated molecular patterns and then evoke immune responses appropriate to the nature of the invading pathogen(s). Because innate immune cells express various types of these receptors, distinct combinations of signaling pathways are activated in response to a given pathogen. Although the conventional wisdom is that these signaling pathways cooperate with one another to ensure an effective host response, a more nuanced view recognizes antagonism between the individual pathways, where the attenuation of a signaling pathway(s) by others may shape the immune response. In this study, we show that, on Listeria monocytogenes infection, Toll-like receptor-triggered MyD88 signaling pathways suppress type I IFN gene induction, which is detrimental to macrophage bactericidal activity. These pathways target and suppress the IFN regulatory factor 3 (IRF3) transcription factor that is activated by the stimulator of IFN genes-TANK-binding kinase-1 kinase pathway. We also provide evidence for the involvement of the MAPK phosphatase family members, which renders IRF3 hypophosphorylated on Toll-like receptor signaling by enhancing the formation of an MAPK phosphatase-IRF3-TANK-binding kinase-1 ternary complex. This study, therefore, reveals a hitherto unrecognized and important contribution of a beneficial innate signaling interference against bacterial infections.

Recent discoveries in signal-transducing innate receptors have illustrated the close link between innate and adaptive immunity. These advances revisit a fundamental issue of immunology, the recognition of self and non-self molecules by the immune system. Indeed, mounting evidence has been provided that the sensing of self-derived molecules by the immune system is important for health and disease. The high-mobility group box (HMGB) proteins, particularly HMGB1, are self-derived immune activators that have multiple functions in the regulation of immunity and inflammation. In this review, we summarize current knowledge of the function of HMGB proteins, as a ligand that can evoke inflammatory responses, and as a sensor for nucleic-acid-mediated immune responses.

Although the mechanisms by which innate pathogen-recognition receptors enhance adaptive immune responses are increasingly well understood, whether signaling events from distinct classes of receptors affect each other in modulating adaptive immunity remains unclear. We found here that the activation of cytosolic RIG-I-like receptors (RLRs) resulted in the selective suppression of transcription of the gene encoding the p40 subunit of interleukin 12 (Il12b) that was effectively induced by the activation of Toll-like receptors (TLRs). The RLR-activated transcription factor IRF3 bound dominantly, relative to IRF5, to the Il12b promoter, where it interfered with the TLR-induced assembly of a productive transcription-factor complex. The activation of RLRs in mice attenuated TLR-induced responses of the T helper type 1 cell (T(H)1 cell) and interleukin 17-producing helper T cell (T(H)17 cell) subset types and, consequently, viral infection of mice caused death at sublethal doses of bacterial infection. The innate immune receptor cross-interference we describe may have implications for infection-associated clinical episodes.

Highlights: {yields} We identified RNA-binding motif protein 3 (RBM3) as CpG-B DNA-binding protein. {yields} RBM3 translocates from the nucleus to the cytoplasm and co-localized with CpG-B DNA. {yields} We newly generated Rbm3-deficient (Rbm3{sup -/-}) mice. {yields} DNA-mediated cytokine gene induction was normally occured in Rbm3{sup -/-} cells. {yields}Rbm3{sup -/-} MEFs showed poorer proliferation rate and increased number of G2-phase cells. -- Abstract: The activation of innate immune responses is critical to host defense against microbial infections, wherein nucleic acid-sensing pattern recognition receptors recognize DNA or RNA from viruses or bacteria and activate downstream signaling pathways. In a search for new DNA-sensing molecules that regulate innate immune responses, we identified RNA-binding motif protein 3 (RBM3), whose role has been implicated in the regulation of cell growth. In this study, we generated Rbm3-deficient (Rbm3{sup -/-}) mice to study the role of RBM3 in immune responses and cell growth. Despite evidence for its interaction with immunogenic DNA in a cell, no overt phenotypic abnormalities were found in cells from Rbm3{sup -/-} mice for the DNA-mediated induction of cytokine genes. Interestingly, however, Rbm3{sup -/-} mouse embryonic fibroblasts (MEFs) showed poorer proliferation rates as compared to control MEFs. Further cell cycle analysis revealed that Rbm3{supmore » -/-} MEFs have markedly increased number of G2-phase cells, suggesting a hitherto unknown role of RBM3 in the G2-phase control. Thus, these mutant mice and cells may provide new tools with which to study the mechanisms underlying the regulation of cell cycle and oncogenesis.« less

The activation of the innate immune responses by DNA exposed within the cytosol has gained much attention and, in this context, several cytosolic DNA sensors have been identified. However, previous studies revealed the operation of redundant and complex mechanisms and it still remains to be clarified how the DNA-mediated evocation of diverse innate immune responses can be achieved. Here we show that two RIG-I-like receptors (RLRs), RIG-I and MDA5, known as cytosolic RNA receptors, nonredundantly function as cytosolic DNA receptors that lead to the selective activation of type I IFN genes. We demonstrate that overexpression of otherwise IFN-inducible RIG-I or MDA5 in IFN signal-deficient cells results in a marked enhancement of type I IFN gene induction upon cytosolic DNA stimulation, while in their absence the induction is impaired. Interestingly, the DNA-mediated induction of other cytokine genes was barely affected by the absence of RLRs. Indeed, unlike the RNA-RLR pathway that activates the transcription factors IRF3 and NF-κB, the DNA-RLR pathway is primarily responsible for the IRF3 activation critical for type I IFN gene transcription, illustrating a deliberate divergence of the DNA signaling pathways. Expectedly, the RLR pathway also contributes to intricate innate immune responses against infection by a DNA virus. Our study may provide insights into the complexity of host defense mechanisms that thwart immune evasion by DNA-containing pathogens.

Apoptosis is a highly regulated process of cell suicide that occurs during development, host defense, and pathophysiology. The transcription factor IFN regulatory factor 5 (IRF5), known to be involved in the activation of innate immune responses, recently has been shown to be critical for DNA damage-induced apoptosis and tumor suppression. Here, we report on a cell-type-specific role of IRF5 in promoting apoptosis upon signaling through the death receptor Fas (CD95/APO-1/TNFRSF6). In particular, we show that mice deficient in the Irf5 gene are resistant to hepatic apoptosis and lethality in response to the in vivo administration of a Fas-activating monoclonal antibody, and that IRF5 is involved in a stage of Fas signaling that precedes the activation of caspase 8 and c-Jun N-terminal kinase (JNK). In addition to hepatocytes, IRF5 is also required for apoptosis in dendritic cells activated by hypomethylated CpG but not in thymocytes and embryonic fibroblasts in vitro . Thus, these findings reveal a cell-type-specific function for IRF5 in the complex regulatory mechanism of death-receptor-induced apoptosis.

Central to innate immunity is the sensing of pathogen-associated molecular patterns by cytosolic and membrane-associated receptors. In particular, DNA is a potent activator of immune responses during infection or tissue damage, and evidence indicates that, in addition to the membrane-associated Toll-like receptor 9, an unidentified cytosolic DNA sensor(s) can activate type I interferon (IFN) and other immune responses. Here we report on a candidate DNA sensor, previously named DLM-1 (also called Z-DNA binding protein 1 (ZBP1)), for which biological function had remained unknown; we now propose the alternative name DAI (DNA-dependent activator of IFN-regulatory factors). The artificial expression of otherwise IFN-inducible DAI (DLM-1/ZBP1) in mouse fibroblasts selectively enhances the DNA-mediated induction of type I IFN and other genes involved in innate immunity. On the other hand, RNA interference of messenger RNA for DAI (DLM-1/ZBP1) in cells inhibits this gene induction programme upon stimulation by DNA from various sources. Moreover, DAI (DLM-1/ZBP1) binds to double-stranded DNA and, by doing so, enhances its association with the IRF3 transcription factor and the TBK1 serine/threonine kinase. These observations underscore an integral role of DAI (DLM-1/ZBP1) in the DNA-mediated activation of innate immune responses, and may offer new insight into the signalling mechanisms underlying DNA-associated antimicrobial immunity and autoimmune disorders.

Viral infection activates host defense mechanisms, including the production of type I interferon (IFN) and the apoptosis of infected cells. We investigated whether these two antiviral responses were differentially regulated in infected cells. We showed that the mitogen-activated protein kinase (MAPK) kinase kinase (MAPKKK) apoptosis signal-regulating kinase 1 (ASK1) was activated in cells by the synthetic double-stranded RNA analog polyinosinic:polycytidylic acid [poly(I:C)] and by RNA viruses, and that ASK1 played an essential role in both the induction of the gene encoding IFN-β (IFNB) and apoptotic cell death. In contrast, we found that the MAPKKK ASK2, a modulator of ASK1 signaling, was essential for ASK1-dependent apoptosis, but not for inducing IFNB expression. Furthermore, genetic deletion of either ASK1 or ASK2 in mice promoted the replication of influenza A virus in the lung. These results indicated that ASK1 and ASK2 are components of the antiviral defense mechanism and suggested that ASK2 acts as a key modulator that promotes apoptosis rather than the type I IFN response. Because ASK2 is selectively present in epithelium-rich tissues, such as the lung, ASK2-dependent apoptosis may contribute to an antiviral defense in tissues with a rapid repair rate in which cells could be readily replaced.

The recognition of microbial components by Toll-like receptors (TLRs) initiates signal transduction pathways, which trigger the expression of a series of target genes. It has been reported that TLR signaling is enhanced by cytokines such as IFN-γ, but the mechanisms underlying this enhancement remain unclear. The MyD88 adaptor, which is essential for signaling by many TLRs, recruits members of the IFN regulatory factor (IRF) family of transcription factors, such as IRF5 and IRF7, to evoke the activation of TLR target genes. In this study we demonstrate that IRF1, which is induced by IFN-γ, also interacts with and is activated by MyD88 upon TLR activation. We provide evidence that MyD88-associated IRF1 migrates into the nucleus more efficiently than non-MyD88-associated IRF1 and that this IRF1 selectively participates in the TLR-dependent gene induction program. The critical role of MyD88-dependent “IRF1 licensing” is underscored by the observation that the induction of a specific gene subset downstream of the TLR–MyD88 pathway, such as IFN-β, inducible NO synthase, and IL-12p35, are impaired inIrf1-deficient cells. Thus, our present study places IRF1 as an additional member participating in MyD88 signaling and provides a mechanistic insight into the enhancement of the TLR-dependent gene induction program by IFN-γ.

Summary Interferons (IFNs) elicit multifaceted effects in host innate defence. Accumulating evidence revealed that not only the first identified Jak-Stat pathway but also other newly found signalling pathways are required for the induction of versatile responses by IFNs. In particular, type I IFNs are inducible by viral infection through the recognition of pathogen-associated molecules by pattern recognition receptors, and the induction of multiple IFN-stimulated genes through the activation of type I IFN signalling confers antiviral and immunomodulatory activities. Any step in this process is often targeted by viruses for their immuno-evasion. The regulatory function of constitutive IFN-α/β signalling has been recognized in terms of its boosting effect on cellular responsiveness in host defence systems. Further comprehensive understanding of IFN signalling may offer a better direction to unravelling the complex signalling networks in the host defence system, and may contribute to their more effective therapeutic applications.

Epidemiological studies have demonstrated that asymptomatic infection of Borna disease virus (BDV) is found in various species of animals in Japan. Recent reports have also revealed that neurological diseases caused by this virus could exist in horses, cattle, a dog, and cats in this country. In this study, we investigated seroprevalence of BDV antibodies in Japanese black cows reared in Kyushu, the southernmost main island of Japan, using ELISA and Western-immunoblotting. Of 101 serum samples, 11 (10.9%) and 21(20.7%) sera were identified as having antibodies to the BDV N and P antigens, respectively. Among the positive sera, three cows (2.9%) were seropositive for both of the antigens. Furthermore, interestingly, only female cows showed antibodies to P, whereas N antibodies were detected in male and female cows with a comparative ratio. Together with previous studies, our results indicate that BDV might be widely spread in cattle raised in Japan. Furthermore, this is the first report to show that beef cattle, Japanese black cattle, have antibodies against a possible zoonotic pathogen, BDV.

Robust type-I interferon (IFN-alpha/beta) induction in plasmacytoid dendritic cells, through the activation of Toll-like receptor 9 (TLR9), constitutes a critical aspect of immunity. It is absolutely dependent on the transcription factor IRF-7, which interacts with and is activated by the adaptor MyD88. How plasmacytoid dendritic cells, but not other cell types (such as conventional dendritic cells), are able to activate the MyD88-IRF-7-dependent IFN induction pathway remains unknown. Here we show that the spatiotemporal regulation of MyD88-IRF-7 signalling is critical for a high-level IFN induction in response to TLR9 activation. The IFN-inducing TLR9 ligand, A/D-type CpG oligodeoxynucleotide (CpG-A), is retained for long periods in the endosomal vesicles of plasmacytoid dendritic cells, together with the MyD88-IRF-7 complex. However, in conventional dendritic cells, CpG-A is rapidly transferred to lysosomal vesicles. We further show that conventional dendritic cells can also mount a robust IFN induction if CpG-A is manipulated for endosomal retention using a cationic lipid. This strategy also allows us to demonstrate endosomal activation of the IFN pathway by the otherwise inactive TLR9 ligand B/K-type oligodeoxynucleotide (CpG-B). Thus, our study offers insights into the regulation of TLR9 signalling in space, potentially suggesting a new avenue for therapeutic intervention.

The activation of Toll-like receptors (TLRs) is central to innate and adaptive immunity. All TLRs use the adaptor MyD88 for signalling, but the mechanisms underlying the MyD88-mediated gene induction programme are as yet not fully understood. Here, we demonstrate that the transcription factor IRF-5 is generally involved downstream of the TLR-MyD88 signalling pathway for gene induction of proinflammatory cytokines, such as interleukin-6 (IL-6), IL-12 and tumour-necrosis factor-alpha. In haematopoietic cells from mice deficient in the Irf5 gene (Irf5-/- mice), the induction of these cytokines by various TLR ligands is severely impaired, whereas interferon-alpha induction is normal. We also provide evidence that IRF-5 interacts with and is activated by MyD88 and TRAF6, and that TLR activation results in the nuclear translocation of IRF-5 to activate cytokine gene transcription. Consistently, Irf5-/- mice show resistance to lethal shock induced by either unmethylated DNA or lipopolysaccharide, which correlates with a marked decrease in the serum levels of proinflammatory cytokines. Thus, our study identifies IRF-5 as a new, principal downstream regulator of the TLR-MyD88 signalling pathway and a potential target of therapeutic intervention to control harmful immune responses.

Toll-like receptor (TLR) activation is central to immunity, wherein the activation of the TLR9 subfamily members TLR9 and TLR7 results in the robust induction of type I IFNs (IFN-α/β) by means of the MyD88 adaptor protein. However, it remains unknown how the TLR signal “input” can be processed through MyD88 to “output” the induction of the IFN genes. Here, we demonstrate that the transcription factor IRF-7 interacts with MyD88 to form a complex in the cytoplasm. We provide evidence that this complex also involves IRAK4 and TRAF6 and provides the foundation for the TLR9-dependent activation of the IFN genes. The complex defined in this study represents an example of how the coupling of the signaling adaptor and effector kinase molecules together with the transcription factor regulate the processing of an extracellular signal to evoke its versatile downstream transcriptional events in a cell. Thus, we propose that this molecular complex may function as a cytoplasmic transductional-transcriptional processor.

A complex mechanism may be operational for dendritic cell (DC) maturation, wherein Toll-like receptor and other signaling pathways may be coordinated differently depending on the nature of the pathogens, in order for DC maturation to be most effective to a given threat. Here, we show that IFN-α/β signaling is selectively required for the maturation of DCs induced by double-stranded RNA or viral infection in vitro . Interestingly, the maturation is still observed in the absence of either of the two target genes of IFN-α/β, TLR3 and PKR (double-stranded-RNA-dependent protein kinase R), indicating the complexity of the IFN-α/β-induced transcriptional program in DCs. We also show that the DCs stimulated in vivo by these agents can migrate into the T cell zone of the spleen but fail to mature without the IFN signal. The immune system may have acquired the selective utilization of this cytokine system, which is essential for innate antiviral immunity, to effectively couple with the induction of adaptive immunity.

The eradication of tumor cells requires communication to and signaling by cells of the immune system. Natural killer (NK) cells are essential tumor-killing effector cells of the innate immune system; however, little is known about whether or how other immune cells recognize tumor cells to assist NK cells. Here, we show that the innate immune receptor Dectin-1 expressed on dendritic cells and macrophages is critical to NK-mediated killing of tumor cells that express N-glycan structures at high levels. Receptor recognition of these tumor cells causes the activation of the IRF5 transcription factor and downstream gene induction for the full-blown tumoricidal activity of NK cells. Consistent with this, we show exacerbated in vivo tumor growth in mice genetically deficient in either Dectin-1 or IRF5. The critical contribution of Dectin-1 in the recognition of and signaling by tumor cells may offer new insight into the anti-tumor immune system with therapeutic implications. DOI: http://dx.doi.org/10.7554/eLife.04177.001, eLife digest When cells in the body grow and divide uncontrollably, cancerous tumors can form. An individual's likelihood of recovering from cancer is highly variable and often depends on the type of cancer and the extent of the disease at the start of treatment. Researchers are therefore interested in discovering how the body responds against cancerous cells. The first line of defense against infection and disease is the body's innate immune system, which includes a suite of immune cells known as white blood cells. These cells patrol the body's organs and tissues in an effort to immediately respond to pathogens and damaged, stressed or otherwise abnormal host cells. Among white blood cells, natural killer cells are involved in identifying and destroying tumor cells. However, it was unclear whether or how other immune cells might help natural killer cells to destroy tumors. In addition, although immune cells detect pathogens and injured cells by producing proteins called pattern recognition receptors, it was unknown whether these receptors also detect tumor cells. Here, Chiba et al. reveal that two other types of immune cell—dendritic cells and macrophages—play essential roles in helping natural killer cells to prevent tumors from growing in mice. The dendritic cells and macrophages produce a pattern recognition receptor called Dectin-1 that recognizes a molecule found on the surface of some—but not all—types of tumor cell. In doing so, Dectin-1 activates a critical signaling pathway and directs the activity of the natural killer cells so that they can effectively kill tumor cells. Chiba et al. found that these tumors grew faster in mice that lack the Dectin-1 protein. The findings of Chiba et al. may also help to explain the effectiveness of certain antibodies—proteins that recognize and neutralize foreign objects such as bacteria and viruses—in cancer therapy. In addition, the Dectin-1 pathway presents a new avenue of research that may offer new cancer treatments. DOI: http://dx.doi.org/10.7554/eLife.04177.002

The large intestinal epithelial cells and immune cells are exposed to a variety of molecules derived from commensal microbiota that can activate innate receptors, such as Toll-like receptors (TLRs) and retinoic acid-inducible gene-I-like receptors (RLRs). Although the activation of these receptors is known to be critical for homeostasis of the large intestine, the underlying gene regulatory mechanisms are not well understood. Here, we show that IFN regulatory factor (IRF)3 is critical for the suppression of dextran sulfate sodium-induced colitis. IRF3-deficient mice exhibited lethal defects in the inflammatory and recovery phases of the colitis, accompanied by marked defects in the gene induction for thymic stromal lymphopoietin (TSLP), a cytokine known to be essential for protection of the large intestine. We further provide evidence that DNA and RNA of the large intestinal contents are critical for Tslp gene induction via IRF3 activation by cytosolic nucleic acid receptors. We also demonstrate that IRF3 indeed activates the gene promoter of Tslp via IRF-binding sequences. This newly identified intestinal gene regulatory mechanism, wherein IRF3 activated by microbiota-derived nucleic acids plays a critical role in intestinal homeostasis, may have clinical implication in colonic inflammatory disorders.

The activation of innate immune responses by nucleic acids is central to the generation of host responses against pathogens; however, nucleic acids can also trigger the development and/or exacerbation of pathogenic responses such as autoimmunity. We previously demonstrated that the selective activation of nucleic acid-sensing cytosolic and Toll-like receptors is contingent on the promiscuous sensing of nucleic acids by high-mobility group box proteins (HMGBs). From this, we reasoned that nonimmunogenic nucleotides with high-affinity HMGB binding may function as suppressing agents for HMGB-mediated diseases, particularly those initiated and/or exacerbated by nucleic acids. Here we characterize an array of HMGB-binding, nonimmunogenic oligodeoxynucleotides (ni-ODNs). Interestingly, we find that binding affinity is rather independent of nucleotide sequence, but is instead dependent on length and structure of the deoxyribose backbone. We further show that these ni-ODNs can strongly suppress the activation of innate immune responses induced by both classes of nucleic acid-sensing receptors. We also provide evidence for the suppressive effect of an ni-ODN, termed ISM ODN, on the induction of adaptive immune responses and in mouse models of sepsis and autoimmunity. We discuss our findings in relation to the critical role of HMGBs in initiating immune responses and the possible use of these ni-ODNs in therapeutic interventions.

Hypertension is a typical modern lifestyle-related disease that is closely associated with the development of cardiovascular disorders. Elevation of angiotensin II (ANG II) is one of several critical factors for hypertension and heart failure; however, the mechanisms underlying the ANG II-mediated pathogenesis are still poorly understood. Here, we show that ANG II-mediated cardiac fibrosis, but not hypertrophy, is regulated by interferon regulatory factor 3 (IRF3), which until now has been exclusively studied in the innate immune system. In a ANG II-infusion mouse model (3.0 mg/kg/d), we compared IRF3-deficient mice (Irf3(-/-)/Bcl2l12(-/-)) with matched wild-type (WT) controls. The development of cardiac fibrosis [3.95 ± 0.62% (WT) vs. 1.41 ± 0.46% (Irf3(-/-)/Bcl2l12(-/-)); P

Interferon regulatory factor (IRF) 5 is a key transcription factor for the activation of innate immune responses downstream of Toll-like receptor signaling. Based on recent genetic analyses, IRF5 is a focus for its potential involvement in systemic lupus erythematosus (SLE), although how IRF5 contributes to SLE is uncertain. In this study, we demonstrate a requirement for IRF5 in the development of murine SLE via its role in B lymphocytes. We show that antinuclear autoantibodies and Ig glomerular deposits, hallmarks of SLE, are absent in Irf5 −/− mice challenged to develop SLE by pristane injection. In particular, production of autoantibodies of the IgG2a subtype, the most prominent isotype in inducing autoimmunity, requires IRF5. Finally, we provide evidence for the critical role of this transcription factor in the secretion of pathogenic antibodies through its direct control of class switch recombination of the γ2a locus. By demonstrating a B-cell-intrinsic role, this study places IRF5 in a context that may have implications for understanding the pathogenesis of human SLE.

The activation of innate immune responses by nucleic acids is crucial to protective and pathological immunities and is mediated by the transmembrane Toll-like receptors (TLRs) and cytosolic receptors. However, it remains unknown whether a mechanism exists that integrates these nucleic-acid-sensing systems. Here we show that high-mobility group box (HMGB) proteins 1, 2 and 3 function as universal sentinels for nucleic acids. HMGBs bind to all immunogenic nucleic acids examined with a correlation between affinity and immunogenic potential. Hmgb1(-/-) and Hmgb2(-/-) mouse cells are defective in type-I interferon and inflammatory cytokine induction by DNA or RNA targeted to activate the cytosolic nucleic-acid-sensing receptors; cells in which the expression of all three HMGBs is suppressed show a more profound defect, accompanied by impaired activation of the transcription factors interferon regulatory factor 3 (IRF3) and nuclear factor (NF)-kappaB. The absence of HMGBs also severely impairs the activation of TLR3, TLR7 and TLR9 by their cognate nucleic acids. Our results therefore indicate a hierarchy in the nucleic-acid-mediated activation of immune responses, wherein the selective activation of nucleic-acid-sensing receptors is contingent on the more promiscuous sensing of nucleic acids by HMGBs. These findings may have implications for understanding the evolution of the innate immune system and for the treatment of immunological disorders.

Nine interferon regulatory factors (IRFs) compose a family of transcription factors in mammals. Although this family was originally identified in the context of the type I interferon system, subsequent studies have revealed much broader functions performed by IRF members in host defense. In this review, we provide an update on the current knowledge of their roles in immune responses, immune cell development, and regulation of oncogenesis.

Interferons-alpha/beta, which are produced upon viral infection, are key soluble factors for the establishment of an antiviral state, but are also produced at low levels in the absence of infection. Herein, we demonstrate that a weak signal by these constitutively produced IFN-alpha/beta show a preventive role in cellular transformation. Ifnar1-deficient (Ifnar1(-/-)) MEF, which are devoid of IFN-alpha/beta signal, undergo a spontaneous transformation during long-term cell culture. Similar to Irf1(-/-) MEF, primary Ifnar1(-/-) MEF become tumorigenic in nude mice by the expression of activated c-Ha-Ras oncoprotein. However, Ifnar1(-/-) MEF do not show any abnormal growth properties. A similar observation is made in Ifnb(-/-) MEF that fail to produce constitutive IFN-alpha/beta, whereas such a transforming property is not found in MEF that lack any of the IFN receptor downstream molecules including Stat1, IRF9 and IRF1. Furthermore, Ifnar1(-/-) mice develop chemically-induced skin papilloma more severely than wild-type mice. In addition, the expression levels of IFNAR1 mRNA are significantly decreased in human gastric cancer tissues. These results suggest a cell-intrinsic role of the weak signal by constitutively produced IFN-alpha/beta to prevent cells from transformation, which may be mediated by a hitherto-unknown pathway(s) downstream of the IFN-alpha/beta receptor.

The sensing of nucleic acids by pattern recognition receptors is a central feature of innate immunity and is mediated by two types of receptors, Toll-like receptors and cytosolic receptors. Indeed, DNA, be it pathogen-derived or self-derived, can potently trigger the innate immune system; and much attention has been focused on the regulation of the DNA-sensing system(s) in the context of protective and pathological immune responses. An accumulating body of evidence has demonstrated that in addition to the membrane-bound type DNA-sensing receptor TLR9, there are cytosolic DNA receptors that can also evoke these responses. In particular, DAI (DLM-1/ZBP1), Trex1, and other regulators of the cytosolic DNA-sensing system have recently been identified and characterized. Here, we summarize our current understanding of how cytosolic DNA receptors contribute to the regulation of innate immune responses and discuss the complexity of the cytosolic DNA-sensing system as well as its future prospects.

A conundrum of innate antiviral immunity is how nucleic acid-sensing Toll-like receptors (TLRs) and RIG-I/MDA5 receptors cooperate during virus infection. The conventional wisdom has been that the activation of these receptor pathways evokes type I IFN (IFN) responses. Here, we provide evidence for a critical role of a Toll-like receptor 3 (TLR3)-dependent type II IFN signaling pathway in antiviral innate immune response against Coxsackievirus group B serotype 3 (CVB3), a member of the positive-stranded RNA virus family picornaviridae and most prevalent virus associated with chronic dilated cardiomyopathy. TLR3-deficient mice show a vulnerability to CVB3, accompanied by acute myocarditis, whereas transgenic expression of TLR3 endows even type I IFN signal-deficient mice resistance to CVB3 and other types of viruses, provided that type II IFN signaling remains intact. Taken together, our results indicate a critical cooperation of the RIG-I/MDA5-type I IFN and the TLR3-type II IFN signaling axes for efficient innate antiviral immune responses.

DNA, whether it is microbe-derived or host-derived, evokes immune responses when exposed to the cytosol of a cell. We previously reported that DNA-dependent activator of IFN regulatory factors (DAI), also referred to as DLM-1/ZBP1, functions as a DNA sensor that activates the innate immune system. In the present study, we examined the regulation of the complex DNA-sensing system by DAI and other molecules. We first show that DAI directly interacts with DNA in vitro and that it requires three DNA-binding domains for full activation in vivo . We also show that the artificially induced dimerization of DAI results in the DNA-independent activation of type I IFN genes, thereby providing a better understanding for the molecular basis of DAI activation. Furthermore, we provide evidence for the presence of additional DNA sensors, either positively or negatively regulating cytosolic DNA-mediated innate immune responses. These results in toto provide insights into the mechanism of DAI activation and reveal the complex regulatory mechanisms underlying DNA-mediated protective and pathologic immune responses.

The interferon regulatory factor (IRF) family, consisting of nine members in mammals, was identified in the late 1980s in the context of research into the type I interferon system. Subsequent studies over the past two decades have revealed the versatile and critical functions performed by this transcription factor family. Indeed, many IRF members play central roles in the cellular differentiation of hematopoietic cells and in the regulation of gene expression in response to pathogen-derived danger signals. In particular, the advances made in understanding the immunobiology of Toll-like and other pattern-recognition receptors have recently generated new momentum for the study of IRFs. Moreover, the role of several IRF family members in the regulation of the cell cycle and apoptosis has important implications for understanding susceptibility to and progression of several cancers.

Borna disease virus (BDV) is a neurotropic virus that causes a persistent infection in the central nervous system (CNS) of many vertebrate species. Although a severe reactive gliosis is observed in experimentally BDV-infected rat brains, little is known about the glial reactions contributing to the viral persistence and immune modulation in the CNS. In this regard, we examined the expression of an astrocyte-derived factor, S100B, in the brains of Lewis rats persistently infected with BDV. S100B is a Ca 2+ -binding protein produced mainly by astrocytes. A prominent role of this protein appears to be the promotion of vascular inflammatory responses through interaction with the receptor for advanced glycation end products (RAGE). Here we show that the expression of S100B is significantly reduced in BDV-infected brains despite severe astrocytosis with increased glial fibrillary acidic protein immunoreactivity. Interestingly, no upregulation of the expression of S100B, or RAGE, was observed in the persistently infected brains even when incited with several inflammatory stimuli, including lipopolysaccharide. In addition, expression of the vascular cell adhesion molecule 1 (VCAM-1), as well as the infiltration of encephalitogenic T cells, was significantly reduced in persistently infected brains in which an experimental autoimmune encephalomyelitis was induced by immunization with myelin-basic protein. Furthermore, we demonstrated that the continuous activation of S100B in the brain may be necessary for the progression of vascular immune responses in neonatally infected rat brains. Our results suggested that BDV infection may impair astrocyte functions via a downregulation of S100B expression, leading to the maintenance of a persistent infection.

Host defense consists of two main aspects, namely, immune response to invading pathogens and suppression of tumor development. A family of transcription factors, IFN regulatory factors (IRFs), has recently gained much attention in terms of its critical role in linking these two aspects of host defense, wherein IRF5 was previously shown to play a critical role in the induction of proinflammatory cytokines by activation of Toll-like receptors. In the present study, using IRF5 gene-targeted mice ( Irf5 −/− mice), we demonstrate another facet of the IRF5 function in the regulation of immune response and tumor suppression. We show that IRF5 is critical for antiviral immunity by showing that Irf5 −/− mice are highly vulnerable to viral infections, accompanied by a decrease in type I IFN induction in the sera. Furthermore, we show that Irf5 −/− fibroblasts are resistant to apoptosis upon viral infection, resulting in an enhanced viral propagation. Finally, we provide evidence that IRF5 is critical for the induction of apoptosis, but not in cell cycle arrest, in response to DNA damage and that IRF5 functions as a tumor suppressor by acting on a pathway that may be distinct from that for p53. These results, together with the dual regulation of IRF5 gene expression by IFN signaling and p53, may provide a new link in the transcriptional network underlying antiviral immunity and tumor suppression.

The recognition of microbial components by Toll-like receptors (TLRs) is an event central to the activation of innate and adaptive immune systems. TLR activation triggers the induction of downstream target genes, wherein the TLR-interacting adaptor molecule MyD88 recruits various signaling molecules and transcription factors. Two members of the IFN regulatory factor (IRF) family of transcription factors, IRF-5 and IRF-7, interact with MyD88 and induce proinflammatory cytokines and type I IFNs, respectively. Here, we show that IRF-4 also interacts with MyD88 and acts as a negative regulator of TLR signaling. IRF-4 mRNA is induced by TLR activation, and IRF-4 competes with IRF-5, but not with IRF-7, for MyD88 interaction. The TLR-dependent induction of proinflammatory cytokines is markedly enhanced in peritoneal macrophages from mice deficient in the Irf4 gene, whereas the induction is inhibited by the ectopic expression of IRF-4 in a macrophage cell line. The critical function of IRF-4 in TLR signaling in vivo is underscored by the observation that Irf4 -deficient mice show hypersensitivity to DNA-induced shock, with elevated serum proinflammatory cytokine levels. This study may provide an insight into the complex regulatory mechanisms of MyD88 signaling by IRFs.

The type I IFN-alpha/beta gene family was identified about a quarter of a century ago as a prototype of many cytokine gene families, which led to the subsequent burst of studies on molecular mechanisms underlying cytokine gene expression and signaling. Although originally discovered for their activity to confer an antiviral state on cells, more evidence has recently been emerging regarding IFN-alpha/beta actions on cell growth, differentiation and many immunoregulatory activities, which are of even greater fundamental biological significance. Indeed, much attention has recently been focused on the induction and function of the IFN-alpha/beta system regulated by Toll-like receptors (TLRs), which are critical for linking the innate and adaptive immunities. The understanding of the regulatory mechanisms of IFN-alpha/beta gene induction by TLRs and viruses is an emerging theme, for which much new insight has been gained over the past few years.